WO2020115021A1

WO2020115021A1 - Method for supporting a diagnosis and device for same

Info

Publication number: WO2020115021A1
Application number: PCT/EP2019/083438
Authority: WO
Inventors: Stephan Felix; Rico FELDTMANN; Andreas KÜMMEL; Bishwas CHAMLING; Doreen BIEDENWEG; Oliver Otto
Original assignee: Universität Greifswald
Priority date: 2018-12-07
Filing date: 2019-12-03
Publication date: 2020-06-11
Also published as: DE102018221232A1

Abstract

The invention relates to a method for supporting a diagnosis as to whether a dilated cardiomyopathy or an inflammatory cardiomyopathy is present, having the following steps: - determining a mechanical property of lymphocytes contained in a blood sample, - comparing the mechanical property with a previously defined reference, and - deciding whether an indicator for dilated cardiomyopathy or inflammatory cardiomyopathy is present on the basis of the comparison.

Description

Diagnostic support method and apparatus therefor

Technical field

The present invention relates to a method for

Supporting a diagnosis of whether there is inflammatory cardiomyopathy. The invention further relates to a device which is designed to carry out such a method.

Technical field

Heart failure is one of the most common

Causes of mortality around the world and is therefore a huge burden on the health care system.

Dilated cardiomyopathy (DCM) is responsible for 30-40% of all heart failure cases and is one of the main reasons for heart transplants. Inflammatory cardiomyopathy

(DCMi) is a common form of DCM. A

longitudinal prospective examination of a larger patient cohort with V. a. Myocarditis showed that the immunohistological evidence of myocardial inflammation in a multivariate analysis was associated with a poor prognosis, whereas a restricted left ventricular pump function was only univarially associated with a poor prognosis. (Kindermann et al. Predictors of outcome in patients with suspected myocarditis. Circulation

2008; 118: 639-648.) Therefore, the detection of myocardial inflammation in the immunohistological analysis of

Myocardial biopsies are significant clinical and

scientific importance. With an inflammatory

Cardiomyopathy play disorders of the cellular immune system along with activation of lymphocytes and

Monocytes / macrophages play an important role. According to a European position paper

Society for Cardiology (ESC) is the dilated

Cardiomyopathy is a disease caused by the presence of left ventricular or biventricular dilatation and systolic dysfunction (left ventricular

Pump function of less than 45%) is defined (Pinto, YM, et al., Proposal for a revised definition of dilated cardiomyopathy, hypokinetic non-dilated cardiomyopathy, and its implications for clinical practice: a Position Statement of the ESC working group on myocardial and pericardial diseases. Eur Heart J, 2016. 37 (23): pp. 1850-8). Large clinical studies have shown that DCM is responsible for many cases of heart failure (HI) and is the main cause of heart transplantation (Haas, J., et al., Atlas of the clinical genetics of human dilated

cardiomyopathy. Eur Heart J, 2015. 36 (18): pp. 1123-35a). The causes of DCM can be both genetic and non-genetic. According to experimental and clinical data, viral infections and myocardial

Inflammatory processes involved in the pathogenesis of DCMi and also DCM (Heymans, S., et al., The Quest for New Approaches in Myocarditis and Inflammatory Cardiomyopathy. J Am Coli Cardiol, 2016. 68 (21): pp. 2348-2364) .

Chronic myocarditis together with ventricular systolic dysfunction is called “inflammatory or

inflammatory cardiomyopathy "(DCMi) and is the most common form of DCM that is not caused by genetic causes (Caforio, AL, et al., Current state of knowledge on aetiology, diagnosis, management, and therapy of myocarditis: a Position Statement of the European Society of Cardiology Working Group on

Myocardial and Pericardial Diseases. Eur Heart J, 2013.

34 (33): pp. 2636-48, 2648a-2648d). Disorders of the cellular and humoral immune system play a causally pathogenically relevant role in the development of DCM (Felix, SB, D. Beug, and M. Dorr, Immunoadsorption therapy in dilated cardiomyopathy. Expert Rev Cardiovasc Ther, 2015. 13 (2): pp. 145-52). In many cases, there will be one

myocardial viral infection induces autoimmune processes that result in activation of the cellular and humoral

Manifest immune system against cardiac antigens. The

Disease course of chronic autoantigen-driven myocardial inflammation can develop into a terminal stage

Cause heart failure. In many cases, the patient can only have one palliative therapy option

Heart transplantation are offered. In patients with

DCHi were suspected in immunohistological methods for the detection of myocardial inflammation

Cardiac muscle samples (endomyocardial biopsies [EMBs]) introduced. The diagnosis of DCMi is based on evidence of an increased number of lymphocytes and macrophages in the

Myocardial tissue as well as an increased expression of

Cell adhesion molecules (Kindermann et al., Caforio et al.).

A fundamental problem in diagnosing patients with DCM is whether DCMi or DCM is of non-inflammatory origin. The clinical pictures are very similar, and it is only possible with great effort, which also requires the removal of EMBs, to distinguish between these two diseases.

It is currently the gold standard for diagnosis

myocardial inflammation an immunohistochemistry of EMBs. There are different statements from the international

Cardiovascular disease societies published on the way of diagnosis. The working group for heart muscle and pericardial diseases of the ESC

recommends in patients with suspected clinical

Myocarditis EMBs for immunohistochemical and molecular

Analyzes. In contrast, the current guidelines for managing heart failure make the American Heart

Association (AHA) and the American College of Cardiology (ACC) contradictory statements regarding the Indication of EMBs for patients with suspected myocarditis. According to the ACC / AHA recommendation, EMBs are only indicated in patients in life-threatening situations, such as unexplained acute cardiomyopathy with inotropic or mechanical circulatory support and / or severe cardiac arrhythmia (Yancy, CW, et al., 2013 ACCF / AHA guideline for the management of heart failure: executive summary: a report of the American College of Cardiology Foundation / American Heart Association Task Force on practice guidelines, Circulation, 2013. 128 (16): pp. 1810-52, Bozkurt, B., et al ., Current Diagnostic and

Treatment Strategies for Specific Dilated Cardiomyopathies: A Scientific Statement From the American Heart Association. Circulation, 2016. 134 (23): pp. E579-e646). These divergent statements by the ESC and the ACC / AHA are examples of the uncertainties with regard to the indication of EMBs in myocarditis. For this reason, they are alternative

Procedures that indicate myocardial inflammation are of particular clinical and scientific interest.

Magnetic resonance imaging for myocarditis as a new imaging method to assess a

Myocarditis (Friedrich, MG, et al., Cardiovascular magnetic resonance in myocarditis: A JACC White Paper. J Am Coli Cardiol, 2009. 53 (17): pp. 1475-87) can be very common in patients with pacemakers or defibrillators) often implanted with this disease, not

be used. Furthermore, the specificity is one

Magnetic resonance imaging for the heart is not high enough to reliably diagnose myocarditis in chronic myocarditis and DCMi. The diagnostic

The gold standard for diagnosing myocarditis is therefore still an immunohistological examination of an EMB (Caforio et al.).

In view of the significant impact on the

Course of the disease when diagnosed with myocarditis Heart failure patients with DCM and the

Contradictory statements from global cardiological societies with regard to the indication of EMBs warrant an intensification of research efforts in order to search for new biomarkers with a

Myocarditis associated with DCM. In addition to the humoral immune system with production of cardiac autoantibodies, the cellular immune system with activation of

Monocytes / macrophages and lymphocytes against cardiac epitopes are involved in the development of myocarditis and consecutive DCMi. In addition to the immunohistochemistry of EMBs, activation of the cellular immune system can also be diagnosed by immunoprofing of peripheral leukocytes. Interestingly, compared to healthy control patients, Thl7 lymphocytes are significantly increased in the peripheral blood of DCMi patients (Myers, JM, et al., Cardiac myosin-Thl7 responses promote heart failure in human myocarditis. JCI Insight, 2016. 1 (9) ). In the

Myocarditis in mice has been shown to improve the

Progression of myocardial disease is dependent on the interaction of CD40-CD40L (Bo, H., L. Zhenhu, and Z. Lijian, Blocking the CD40-CD40L interaction by CD40-Ig reduces disease progress in murine myocarditis induced by CVB3.

Cardiovasc Pathol, 2010. 19 (6): pp. 371-6). This complex is one of the most important factors in the activation of B lymphocytes by T lymphocytes (Parker, DC, The functions of antigen recognition in T cell-dependent B cell activation. Semin Immunol, 1993. 5 (6): p. 413 -20).

It could also be shown that stronger antigen signals induce an increased differentiation of IL-17-producing T lymphocytes through communication between CD40 and CD40L (Iezzi, G., et al., CD40-CD40L cross-talk

integrates strong antigenic Signals and microbial Stimuli to induce development of IL-17-producing CD4 + T cells. Proc Natl Acad Sei USA, 2009. 106 (3): pp. 876-81). These results of immunoprofiling are initially for the Basic research of interest, currently of little relevance for clinical practice due to high costs and the lack of high-throughput processes.

BNP (natriuretic peptide type B) and NT-proBNP (English "N-terminal prohormone of brain natriuretic peptide") are also used as diagnostic biomarkers for the

Heart failure used. However, the significance of these markers is what the diagnosis of the etiology

Heart failure concerned, low (Ponikowski, P., et al., 2016 ESC Guidelines for the diagnosis and treatment of acute and chronic heart failure: The Task Force for the diagnosis and treatment of acute and chronic heart failure of the European Society of Cardiology ( ESC) Developed with the special contribution of the Heart Failure Association (HFA) of the ESC. Eur Heart J, 2016. 37 (27): pp. 2129-2200, and Felker, GM, et al., Serial high sensitivity cardiac troponin T measurement in acute heart failure: insights from the RELAX-AHF study. Eur J Heart Fail, 2015. 17 (12): pp. 1262-70).

Presentation of the invention

For the above reasons, it is clear that it is from

It is of considerable relevance to find new methods or markers that facilitate the diagnosis of a DCMi.

According to the inventors' first research results, the mechanical properties of leukocytes are an indicator for monitoring the progress of diseases in inflammation (Toepfner, N., et al., Detection of human disease conditions by single-cell morpho-rheological phenotyping of blood.

Elife, 2018, 7, Otto, 0., et al. , Real-time deformability cytometry: on-the-fly cell mechanical phenotyping. Nat

Methods, 2015. 12 (3): pp. 199-202).

If you take these properties as a measure of the deformation of a cell under an external load and change this Dimension defined with time, represent the mechanical

Properties an indicator ready, which represents the state of the cell and its function.

These properties have increased in recent years

Get relevance to study the interactions of the metabolism and the molecular structure of proteins

(Münder, M.C., et al., A pH-driven transition of the

cytoplasm from a fluid- to a solid-like state promotes entry into dormancy. Elife, 2016, 5) to stem cells

identify (Xavier, M., et al., Mechanical phenotyping of primary human skeletal stem cells in heterogeneous

populations by real-time deformability cytometry. Integr Biol (Camb), 2016. 8 (5): pp. 616-23) or to characterize disease states (Gossett, D.R., et al., Hydrodynamic

Stretching of single cells for large population mechanical phenotyping. Proc Natl Acad Sei U S A, 2012. 109 (20): S.

7630-5).

However, although an initial study of changes in the cell stiffness of cardiac fibroblasts obtained from EMBs using atomic force microscopy has been promising (Glaubitz, M., et al., Stiffness of left ventricular cardiac fibroblasts is associated with ventricular dilation in patients with recent-onset

nonischemic and nonvalvular cardiomyopathy. Circ J, 2014. 78 (7): pp. 1693-700) - the mechanical properties of cells as a clinically relevant parameter in the

Cardiovascular research has not yet been considered and, in particular, it has not been considered to use these properties to diagnose DCMi.

The inventors are of the opinion that the introduction of real-time deformability cytometry (RT-DC) means that

mechanical properties of cells can now be analyzed in real time at rates of up to 1000 cells per second, which is a prerequisite for the development of a new biomarkers (Golfier, S., et al., High-throughput cell mechanical phenotyping for label-free titration assays of cytoskeletal modifications. Cytoskeleton (Hoboken), 2017. 74 (8): pp. 283-296).

The inventors have therefore hypothesized to take the mechanical properties of lymphocytes as a marker for the presence of inflammatory cardiomyopathy. As will become clear from the further explanations, such a marker is suitable for distinguishing from other clinical pictures and could be used to provide a doctor with an additional new diagnostic method.

The invention is defined by claim 1. Preferred embodiments of the invention are defined in the dependent claims. The invention is further defined by the device claim.

A method according to the invention is a computer-aided method to support the diagnosis of inflammatory cardiomyopathy. The fact that a diagnosis is only “supported” is understood to mean that the claimed method cannot or should not serve as the sole diagnosis. In particular, other diagnostic methods can be used to make a final diagnosis. The present method only serves one purpose Doctor or a person who wants to make a diagnosis

Provide (another) means to diagnose inflammatory cardiomyopathy and to distinguish it from other clinical pictures (such as DCM without myocarditis). This method could also be used to monitor the course of myocardial inflammation in patients with DCMi clearly diagnosed in EMBs.

There is a mechanical property of lymphocytes in a blood sample from a patient who is to be diagnosed with DCMi. The size of the lymphocytes can be distinguished from other components of the blood. The mechanical

Properties of these lymphocytes can be determined by a

The device can be determined, for example, by directly exerting a force through it (for example using an atomic force microscope). However, it is also possible in principle and is also preferred in this patent application to deform such cells by means of a flow cytometer, such as the AcCellerator® from Zellmechanik Dresden GmbH, and to measure the mechanical property of the lymphocytes in this way.

The measured mechanical property is compared with a predetermined reference by means of a computer. For example, it can be checked whether the lymphocytes are firmer or less firm than a reference value. On the

Based on this comparison, the computer then decides whether an indicator of an inflammatory

Cardiomyopathy is present.

As will become clear from the further description, the mechanical properties are a good marker for distinguishing between non-inflammatory cardiomyopathy and inflammatory cardiomyopathy.

Such a marker can be obtained easily and in particular less invasively than with EMBs. In principle, organ biopsies pose a risk, albeit slight, of

Patients. In the case of an EMB, patients are through

Cardiac arrhythmias, bleeding or, in the worst case, endangered by perforation of the heart chamber wall. These complications are rare. In principle, however, the diagnostic benefit against the

Risk must be weighed. In this respect it is one

To determine biomarkers safely in most patients, since only blood collection is required, but none further invasive interventions. This could be important not only for the comfort of the patient, but also for the prognosis if this biomarker is clearly associated with a DCMi. In particular, such an intervention is also possible if an EMB would endanger the patient due to a poor general condition.

In the present case, a doctor will use the method according to the invention when he has to diagnose a DCMi or a DCM non-inflammatory genesis in a patient. I.e. the doctor will analyze a blood sample using the claimed method for a patient whose symptoms suggest a suspected DCM or diagnosis of DCM. This analysis then gives the doctor an indication of whether there is a DCMi or whether there is a non-inflammatory DCM.

It is particularly important that the mechanical

Property is determined by measuring the lymphocytes due to the deformation caused by a fluid flow. Such a deformation allows controlled forces to be exerted on the lymphocytes. In particular, if the liquid flow is laminar, the forces can be accurate and

can be reliably quantified, which would not be the case with a turbulent liquid flow or would only be possible to a limited extent. This also makes the process very good

automate because there are many in the liquid flow

Can examine lymphocytes that are transported through the fluid flow as if on a "conveyor belt".

The deformation is preferably measured optically, for example by means of a microscope. Such an optical

Measurement is easy to implement, also because there are various programming libraries and state of the art

Software packages for the evaluation of electronically received images. Such a measurement can be done in real time, i.e. while the images are being created. Hence the

The term real-time deformability cytometry is coined, as described in WO 2015/024690 A1. The relevance of RT-DC in the life sciences has already been shown in basic research and in

applied cell biological research. Under

Using yeast as a model system, it could be shown that a change in the pH value within the cell leads to a phase transition within the cytoplasm from liquid to solid, which leads to a resting state of the cell (see Münder et al.). This observation is very important, for example, to understand the complex interaction of

Understand cell function, metabolism and mechanics.

In addition, RT-DC was also used to identify a mechanically separate subpopulation of mesenchymal stem cells that were not using traditional methods

was representable (see Xavier et al.).

In contrast, immunofluorescence analysis using FACS is a very complex and expensive method compared to RT-DC. Despite extensive staining and classification of different immune cell populations, no significant differences between DCM and DCMi patients could be found. The FACS analysis can be used to explain molecular mechanisms that are responsible for the fact that immune cells have different deformabilities in DCM and DCMi patients. For the clinical

Routine use, the FACS analysis is clearly inferior to the RT-DC in addition to the costs and effort mentioned, also in handling. Furthermore, standard parameters have also shown no significant difference between DCM and DCMi:

An indicator of inflammatory cardiomyopathy is when there is a mechanical property of the cell

is determined which corresponds to a greater deformability than a limit value. In other words, the cell is then "softer" than a threshold value. The decision that there is no indicator of inflammatory cardiomyopathy is made with a mechanical property that one

less deformability than the limit value, i.e. the cell is then "harder" than a threshold value. Thus, by determining the deformability of the cell

determine whether there is an indicator of inflammatory cardiomyopathy.

Such deformability can, for example, by the

Deformation of lymphocytes can be measured by increasing forces and calculated using the modulus of elasticity. It is sufficient to simply use a computer to check whether a cell is firmer or less firm than a certain limit. Such a process has proven to be particularly meaningful.

According to the invention there is also a device which is designed for carrying out a method according to one of the preceding claims. Such a device can in particular be a modified device according to WO

2015/024690 Al. Such a device can preferably display or record the result of the method optically, acoustically and / or by recording the result on a storage medium.

Brief description of the drawings

Figures la to c show a procedure for

Real-time deformability cytometry. Figure ld shows a change in the deformation of

Granulocytes.

Figure 2 shows results of mechanical profiling of lymphocytes.

Detailed description of the pictures

As shown in Figure la, the experimental setup consists of an inverted microscope, in which blood is pumped through a constriction of a microfluidic channel that is arranged on a microscope stage. As the blood passes through a detection chamber, the cells within the channel are deformed by a hydrodynamic interaction, which is recorded by a high-speed video camera.

An example of such a deformation is shown in Figure 1b, which shows a time series of an initially round cell that undergoes a shape change that is essentially similar to a bullet. With that of Otto et al.

algorithm described are the cell size and the

Deformation analyzed in real time. Here, the deformation d is defined as:

where A describes the area of the cell in the image and 1 the length of the cell perimeter in the image.

Since RT-DC is able to measure heterogeneous samples, a mechanical phenotype of whole blood can be obtained without having to specially prepare the sample (see Figure lc). This figure shows a mechanical phenotype of whole blood that shows red blood cells (highest deformation), platelets and leukocytes (bottom). The section shows leukocytes after the separation. The populations of lymphocytes (T cells and B cells) and granulocytes / monocytes were separated by negative magnetic cell sorting.

Figure 1d shows the in vitro stimulation of granulocytes (gran) by phorbol-12-myristat-13-acetate (PMA), which leads to a clear change in the deformation.

The ability of RT-DC to analyze heterogeneous samples makes it possible to measure cells in whole blood directly, without having to clean them beforehand or otherwise

to have to change. Special analysis methods should make it possible to carry out a direct functional analysis, for example of lymphocytes, regardless of cell activation with potential changes.

As shown in Figure 1d, the activation of granulocytes in vitro with PMA resulted in an increased

Deformation of cells (see Golfier et al. And Otto et al).

Previous in vitro studies of patients with

Respiratory diseases show a higher deformation value for neutrophils and monocytes compared to a healthy control group, while a viral infection with the

Epstein-Barr virus also led to an increased deformation of the lymphocyte population in peripheral blood (Toepfner et al.). These data indicate a differential immune cell response to bacterial and viral infection that is of potential diagnostic interest.

With a view to applying the analysis of the mechanical properties of lymphocytes, a first pilot study was carried out in which data from patients with DCM or DCMi and a left ventricular pump function of less than 45% were analyzed. The results of a pilot study are presented in the following table, in which patients with DCM / DCMi were compared with patients with rheumatoid arthritis (RA) and a healthy control group:

As recommended by the ESC, EMBs were taken from all patients for clinical reasons to determine the etiology of the patient

Myocardial Disease Using Immunohistochemistry

diagnose. In addition, a mechanoprofing of the lymphocytes from peripheral blood samples was performed using RT-DC

and the results were compared to those of healthy individuals. Patients with a

rheumatoid arthritis (RA) served as a patient cohort with a systemic inflammatory disease. In addition, immunoprofhing was carried out in all individuals using a FACS analysis of peripheral blood cells.

For the RT-DC of the peripheral leukocytes were 25 mΐ

Citrate blood analyzed at different flow rates. The different flow rates were needed to get cells

exposed to different hydrodynamic forces that lead to different deformations. This approach enables the mechanical characterization of cells regardless of their initial shape. In the current pilot study, only leukocytes in whole blood were selectively analyzed. Figure 2a shows as examples the mechanical profile of peripheral leukocytes of a healthy participant (control), a DCM patient, a DCMi patient and an RA patient after a continuous measurement for 10 minutes each. Two populations are visible here. While

Lymphocytes have a peak with an average cell size of 40 ym ² , for granulocytes / monocytes there is a peak at approximately 60 ym ² . Because lymphocytes play a critical role in DCMi, the inventors focused on analyzing the lymphocyte population by determining a relative change in mean deformation as a function of flow rate. A linear fit of this data gives a deformation index Di, which is the slope of the

Describes the function and that between the

Control group, the DCM group, the DCMi group and the RA group is compared. The results are shown in Figure 2b.

As shown in the left part of Figure 2b, the control group had a mean Di ± standard deviation of 0.14 ± 0.007, while the DCM and DCMi groups respectively increased values of 0.17 ± 0.009 and 0.21 ± 0.012 exhibit.

The comparisons of the pairs lead to statistically significant differences in the Di of the DCM group compared to the

Control group (p = 0.004), the DCMi group compared to the control group (p = 0.001) and the DCMi group compared to the DCM group (p = 0.03). It is interesting to observe an increased Di in patients with DCMi, which is in line with the previous results of an increased deformation of leukocytes in people with respiratory infections

(Toepfner et al.).

In addition, lymphocytes from patients with rheumatoid arthritis were analyzed to determine the mechanical

Compare the cell profile of DCM patients with that of patients with systemic inflammatory disease. The results are shown in the right part of Figure 2b, where the Di of the RA group exceeds that of the DCM group (p = 0.002).

In order to assess the significance of mechanoprofiling for the diagnosis of whether a patient has DCMi, the sensitivity and specificity for all Di limit values were calculated. An optimal limit of Di = 0.194 with a sensitivity and specificity of 0.78 was found, while the receiver operating characteristic showed an area under the curve of 0.83 for both groups. These

Results show that Di as a marker has great potential for the correct diagnosis of DCMi patients.

The abundance of peripheral lymphocyte subpopulations of DCM / DCMi patients was then quantified using FACS analysis as a first step to immunoprofiling

(Esfandiarei, M. and BM McManus, Molecular biology and pathogenesis of viral myocarditis. Annu Rev Pathol, 2008. 3: pp. 127-55). All of these measurements were performed using a Becton Dickinson LSR II using a group of nine fluorochrome-labeled antibodies, and data analysis was performed using the FlowJo software. The cell numbers were normalized to 100,000 CD 45 ⁺ leukocytes (mean value [minimum value, maximum value]). There were significantly increased values of CD4 ⁺ T cells in DCM patients (mean = 9229 [1478, 17.724]; p = 0.007) and DCMi patients (mean = 4823 [486, 8512], p = 0.009) compared to healthy

Control groups (mean = 1596 [42, 2684]) were found.

Similarly, significantly increased levels of CD19 ⁺ B cells and CD8 ⁺ T cells were found in both DCM (CD19 ⁺ mean = 1964 [163, 3663], p = 0.006), (CD8 ⁺ mean = 4729 [1352, 9377], p =

0.018) and DCMi patients (CD19 ⁺ mean = 2665 [731, 5049], p = 0.003), (CD8 ⁺ mean = 4434 [2114, 10.599], p = 0.023) were found in comparison to healthy individuals (CD19 ⁺ B - cell mean = 596 [73, 1360]), (CD8 ⁺ T cells

Mean = 1627, [237, 3160]). However, the number has increased of B and T cells did not differentiate between DCM and DCMi patients. Despite complex coloring and

It is not possible to differentiate between DCM and DCMi using FACS. So that's it

Mechanoprofiling using RT-DC superior to immunoprofiling using FACS.

The available data show for the first time that the

Mechanoprofiling of peripheral lymphocytes can contribute to the diagnosis of DCMi. Interestingly, too

Lymphocytes from patients with RA showed increased cell deformation comparable to the result obtained from patients with DCMi. Therefore, based on the

Mechanoprofiling DCMi cannot be distinguished from an RA, which is an example of a systemic

Is inflammatory disease. However, a distinction between DCMi and RA is not relevant for clinical practice, since in cases of clinical suspicion of cardiac

Manifestation of a systemic immune disease

International guidelines clearly recommend EMBs.

It should also be noted that in the vast majority

Cases (more than 95%) myocarditis or DCMi is not associated with other immune-mediated diseases.

Diagnostic procedures that identify isolated myocarditis can be classified as clinically relevant if they have a poorer prognosis

accompanied .

Mechanoprofiling is therefore believed to be a new method of screening patients with DCMi. The data from the pilot study also show that the

conventional FACS analysis of peripheral blood cells alone is unsuitable to distinguish patients with DCMi from patients with DCM without myocarditis. In this respect, mechanoprofiling appears to be a new and promising strategy for use in routine clinical practice. However, extensive immunoprofiling will be necessary to Link activation states of the lymphocytes with their mechanical properties.

Claims

Expectations

1. Procedure to support a decision whether inflammatory cardiomyopathy or non-inflammatory

Cardiomyopathy is present, with the following steps:

- Determine a mechanical property of

Lymphocytes contained in a blood sample

- comparing the mechanical property with a defined, predetermined reference, and

- Decide based on the comparison whether there is an indicator of inflammatory cardiomyopathy.

2. The method according to any one of the preceding claims, wherein the mechanical property is determined by measuring the deformation of the lymphocytes due to a liquid flow, the measurement preferably being carried out optically, the measurement being more preferably by means of a “real time”

Deformability Cytometry "takes place.

3. The method according to any one of the preceding claims, wherein the decision that there is no indicator of inflammatory cardiomyopathy is made with a mechanical property that is less deformable than a

Limit value, and the decision that a

There is an indicator of inflammatory cardiomyopathy with a mechanical property that is more deformable than the limit value.

4. Device which is designed for performing a method according to one of the preceding claims.